U.S. patent application number 09/171740 was filed with the patent office on 2002-04-11 for sustained release ionic conjugate.
Invention is credited to IGNATIOUS, FRANCIS XAVIER, LOUGHMAN, THOMAS CIARAN, SHALABY, SHALABY WAHBA, TOURAUD, FRANCK-JEAN-CLAUDE.
Application Number | 20020041893 09/171740 |
Document ID | / |
Family ID | 11041150 |
Filed Date | 2002-04-11 |
United States Patent
Application |
20020041893 |
Kind Code |
A1 |
IGNATIOUS, FRANCIS XAVIER ;
et al. |
April 11, 2002 |
SUSTAINED RELEASE IONIC CONJUGATE
Abstract
A method of spherifying a sustained release ionic conjugate
which contains a free carboxyl group-containing biodegradable
polymer and a free amino group-containing drug which are ionically
bonded to each other.
Inventors: |
IGNATIOUS, FRANCIS XAVIER;
(MASSACHUSETTS, MA) ; LOUGHMAN, THOMAS CIARAN;
(DUBLIN, IE) ; SHALABY, SHALABY WAHBA; (PENDLETON,
SC) ; TOURAUD, FRANCK-JEAN-CLAUDE; (VERNON,
FR) |
Correspondence
Address: |
BRIAN R. MORRILL, ESQ.
BIOMEASURE INC
27 MAPLE STREET
MILFORD
MA
01757-3650
US
|
Family ID: |
11041150 |
Appl. No.: |
09/171740 |
Filed: |
April 20, 1999 |
PCT Filed: |
April 22, 1997 |
PCT NO: |
PCT/IE97/00030 |
Current U.S.
Class: |
424/426 ;
424/484 |
Current CPC
Class: |
Y10T 428/2985 20150115;
A61K 38/08 20130101; A61K 9/167 20130101; A61K 9/1647 20130101;
A61K 47/593 20170801; A61K 9/1641 20130101; A61K 47/6927 20170801;
A61K 38/31 20130101; A61K 9/1694 20130101 |
Class at
Publication: |
424/426 ;
424/484 |
International
Class: |
A61F 002/00; A61K
009/14 |
Foreign Application Data
Date |
Code |
Application Number |
Apr 23, 1996 |
IE |
960308 |
Claims
1. A method of making microparticles of a sustained release ionic
conjugate containing a free carboxyl group-containing biodegradable
polymer and a free amino group-containing drug which are ionically
bonded to each other, the method comprising: obtaining a first
solution in which said conjugate is dissolved, wherein said
solution comprises acetone, acetonitrile, ethyl acetate,
tetrahydrofuran, or glyme; mixing said first solution with a first
liquid to form a first dispersion, wherein said first liquid is
miscible with said first solution, and said conjugate is not
soluble in said first liquid and precipitates out of said first
dispersion; and isolating said conjugate from said first
dispersion.
2. A method according to claim 1, wherein said first solution is
added to said first liquid as small droplets.
3. A method according to claim 2, wherein said first solution is
added to said first liquid through an atomizing nozzle.
4. A method according to any one of claims 1-3, wherein said drug
is a peptide.
5. A method according to any preceding claim, wherein said
biodegradable polymer is a polyester made of lactic acid,
e-caprolic acid, glycolic acid, trimethylene carbonate, or
p-dioxanone; or a copolymer thereof.
6. A method according to any preceding claim, wherein said drug is
soluble in said first liquid.
7. A method according to any preceding claim, wherein said
biodegradable polymer is a polyester comprising lactic acid, or
glycolic acid; or a copolymer thereof.
8. A method according to claim 7, wherein said polyester further
contains malic acid, tartaric acid, citric acid, succinic acid, or
glutaric acid.
9. A method according to any one of claims 4-8, wherein said
peptide is a somatostatin or LHRH.
10. A method according to any preceding claim, wherein said first
liquid is alcohol or water; or a mixture thereof.
11. A method according to claim 10, wherein said first liquid is
ethanol maintained between about 0.degree. C. and -30.degree. C. or
isopropyl alcohol maintained between about 0.degree. C. and
-70.degree. C.
12. A method according to any preceding claim, wherein said first
solution contains acetone or acetonitrile.
13. A method according to any preceding claim, wherein said first
solution is obtained by: dissolving said biodegradable polymer in a
second liquid to form a second solution; dissolving said drug in a
third liquid to form a third solution, wherein said third liquid is
miscible with said first liquid and said second liquid; and mixing
said second solution and said third solution to form said first
solution, wherein said mixing causes said drug to ionically bond to
said biodegradable polymer and form said conjugate in said first
solution.
14. A method according to claim 13, wherein NaOH or KOH is added to
the second solution prior to mixing said second solution and said
third solution.
15. A method according to claim 13 or 14, wherein said second
liquid is acetone; and said third liquid is water or acetone; or a
mixture thereof.
16. A method according to any of claims 1-12, wherein said first
solution is obtained by dissolving said biodegradable polymer and
said drug in a second liquid to form said first solution, thereby
forming said conjugate in said first solution.
17. A method according to claim 16, wherein said second liquid is
acetone or a mixture of acetone and water.
18. A method according to claim 17, wherein said biodegradable
polymer is first dissolved in said second liquid, a base is then
added to said second solution, and said drug is subsequently
dissolved in said second liquid.
19. A method according to any preceding claim, wherein said
conjugate is isolated by centrifuging or filtering said first
dispersion.
20. A method according to claim 19, wherein said first solution is
partially or completely evaporated from said first dispersion prior
to isolation of said conjugate.
21. A method according to claim 20, wherein said isolated conjugate
is mixed with an aqueous mannitol solution prior to vacuum
drying.
22. A method of spherifying a sustained release ionic conjugate
comprising a biodegradable polymer containing a free carboxyl
group-containing biodegradable polymer and a free amino
group-containing drug which are ionically bonded to each other,
said method comprising: mixing said conjugate with a first liquid
to form a first dispersion, wherein said conjugate has the shape of
a microparticle and is not soluble in said first liquid; heating
said first dispersion to a temperature greater than the Tg or Tm of
said conjugate; cooling said first dispersion below the Tg or Tm of
said conjugate; mixing said first dispersion with a second liquid
to form a second disperson, wherein said second liquid is miscible
with said first liquid and said conjugate is not soluble in said
second liquid; and isolating said conjugate from said second
dispersion.
23. A method according to claim 22, wherein said conjugate has the
shape of a microcapsule which has an average diameter of between 5
.mu.m to 200 .mu.m prior to mixing with said first liquid and said
first dispersion is stirred prior to said heating or cooling.
24. A method according to claim 22 or 23, wherein said
biodegradable polymer is a polyester made of lactic acid or
glycolic acid; or a copolymer thereof.
25. A method according to any one of claims 22-24, wherein said
drug is a peptide.
26. A method according to any one of claims 22-25, wherein said
first liquid is an oil and said second liquid is hexane.
27. A method according to any one of claims 22-26, further
comprising: rinsing said isolated conjugate with said second
liquid; and vacuum drying said rinsed conjugate.
28. A method according to claim 27, wherein said isolated conjugate
is mixed with an aqueous mannitol solution prior to vacuum
drying.
29. A method of spherifying a sustained release ionic conjugate
containing a free carboxyl group-containing biodegradable polymer
and a free amino group-containing drug which are ionically bonded
to each other, said method comprising: dissolving said conjugate in
a first liquid to form a first solution; stirring said first
solution with a second liquid to form a first dispersion, wherein
said second liquid is immiscible with said first solution;
evaporating said first liquid from said first dispersion to
precipitate said conjugate from said first dispersion; and
isolating said precipitate conjugate from said first
dispersion.
30. A method according to claim 29, wherein said first solution is
added to said second liquid as small droplets.
31. A method according to claim 29 or 30, wherein said first liquid
is acetonitrile and said second liquid is an oil.
32. A method according to claim 31, wherein said oil is silicon
oil, mineral oil, sesame oil, or a vegetable oil.
33. A method according to any one of claims 29-32, wherein said
biodegradable polymer is a polyester comprising lactic acid or
glycolic acid; or a copolymer thereof.
34. A method according to any one of claims 29-33, wherein said
drug is a peptide.
35. A method according to any one of claims 29-34, further
comprising rinsing said isolated conjugate with a third liquid
which is miscible with said second liquid and not a solvent for
said isolated conjugate.
36. A-method according to claim 35, wherein said third liquid is
hexane, heptane, or octane.
37. A method according to any one of claims 29-36, wherein said
isolated conjugate is mixed with an aqueous mannitol solution prior
to vacuum drying.
38. A method of making microparticles of a sustained release ionic
conjugate, substantially as hereinbefore described and
exemplified.
39. A method of spherifying a sustained release ionic conjugate,
substantially as hereinbefore described and exemplified.
40. A method according to claim 8, wherein said polyester comprises
lactic acid, glycolic acid and tartaric acid.
41. Biodegradable polymer comprising lactic acid, .epsilon.-caproic
acid, glycolic acid, trimethylene carbonate, p-dioxanone or a
copolymer thereof and tartaric acid.
42. Microparticles comprising a biodegradable polymer according to
claim 41.
43. Microparticles of a sustained release ionic conjugate
comprising the biodegradable polymer according to claim 41 and a
drug containing one or more free amino groups, wherein the polymer
and drug are ionically bonded.
44. Microparticles according to claim 43 wherein said drug is
selected from the group consisting of growth hormone releasing
peptide luteinizing hormone-releasing hormone, adrenomedullin,
growth hormone, somatostatin, bombesin, gastrin releasing peptide,
calcitonin, bradykinin, galanin, melanocyte stimulating hormone,
growth hormone releasing factor, amylin, tachykinins, secretin,
parathyroid hormone, enkephalin, endothelin, calcitonin gene
releasing peptide, neuromedins, parathyroid hormone related
protein, glucagon, neurotensin, adrenocorticotrophic hormone,
peptide YY, glucagon releasing peptide, vasoactive intestinal
peptide, pituitary adenylated cyclase activating peptide, motilin,
substance P, neuropeptide Y and TSH or an analogue or a fragment
thereof.
45. Microparticles according to claim 44 wherein said drug is
somatostatin or LHRH or an analogue or a fragment thereof.
46. Microparticles according to claim 45 wherein said somatostatin
analogue is D-.beta.-Nal-c[Cys-Tyr-D-Trp-Val-Cys]-Thr-Nh.sub.2.
47. The biodegradable polymer according to claim 41 comprising
lactic acid glycolic acid and tartaric acid.
48. Microparticles comprising a biodegradable polymer according to
claim 47.
49. Microparticles of a sustained release ionic conjugate
comprising the biodegradable polymer according to claim 47 and a
drug containing one or more free amino groups, wherein the polymer
and drug are ionically bonded.
50. Microparticles according to claim 49 wherein said drug is
selected from the group consisting of growth hormone releasing
peptide, luteinizing hormone-releasing hormone, adrenomedullin,
growth hormone, somatostatin, bombesin, gastrin releasing peptide,
calcitonin, bradykinin, galanin, melanocyte stimulating hormone,
growth hormone releasing factor, amylin, tachykinins, secretin,
parathyroid hormone, enkephalin, endothelin, calcitonin gene
releasing peptide, neuromedins, parathyroid hormone related
protein, glucagon, neurotensin, adrenocorticotrophic hormone,
peptide YY, glucagon releasing peptide, vasoactive intestinal
peptide, pituitary adenylated cyclase activating peptide, motilin,
substance P, neuropeptide Y and TSH or an analogue or a fragment
thereof.
51. Microparticles according to claim 50 wherein said drug is
somatostatin or LHRH or an analogue or a fragment thereof.
52. Microparticles according to claim 51 wherein said somatostatin
analogue is D-.beta.-Nal-c[Cys-Tyr-D-Trp-Val-Cys]-Thr-NH.sub.2.
53. The biodegradable polymer according to claim 47 wherein the
ratio of lactic acid to glycolic acid to taratic acid is about 66
to about 33 to about 1, respectively.
54. Microparticles comprising a biodegradable polymer according to
claim 53.
55. Microparticles of a sustained release ionic conjugate
comprising the biodegradable polymer according to claim 53 and a
drug containing one or more free amino groups, wherein the polymer
and drug are ionically bonded.
56. Microparticles according to claim 55 wherein said drug is
selected from the group consisting of growth hormone releasing
peptide, luteinizing hormone-releasing hormone, adrenomedullin,
growth hormone, somatostatin, bombesin, gastrin releasing peptide,
calcitonin, bradykinin, galanin, melanocyte stimulating hormone,
growth hormone releasing factor, amylin, tachykinins, secretin,
parathyroid hormone, enkephalin, endothelin, calcitonin gene
releasing peptide, neuromedins, parathyroid hormone related
protein, glucagon, neurotensin, adrenocorticotrophic hormone,
peptide YY, glucagon releasing peptide, vasoactive intestinal
peptide, pituitary adenylated cyclase activating peptide, motilin,
substance P, neuropeptide Y and TSH or an analogue or a fragment
thereof.
57. Microparticles according to claim 56 wherein said drug is
somatostatin or LHRH or an analogue or a fragment thereof.
58. Microparticles according to claim 57 wherein said somatostatin
analogue is D-.beta.-Nal-c[Cys-Tyr-D-Trp-Val-Cys]-Thr-NH.sub.2.
59. The biodegradable polymer according to claim 47 wherein the
ratio of lactic acid to glycolic acid to taratic acid is about 66
to about 32 to about 2, respectively.
60. Microparticles comprising a biodegradable polymer according to
claim 59.
61. Microparticles of a sustained release ionic conjugate
comprising the biodegradable polymer according to claim 60 and a
drug containing one or more free amino groups, wherein the polymer
and drug are ionically bonded.
62. Microparticles according to claim 61 wherein said drug is
selected from the group consisting of growth hormone releasing
peptide, luteinizing hormone-releasing hormone, adrenomedullin,
growth hormone, somatostatin, bombesin, gastrin releasing peptide,
calcitonin, bradykinin, galanin, melanocyte stimulating hormone,
growth hormone releasing factor, amylin, tachykinins, secretin,
parathyroid hormone, enkephalin, endothelin, calcitonin gene
releasing peptide, neuromedins, parathyroid hormone related
protein, glucagon, neurotensin, adrenocorticotrophic hormone,
peptide YY, glucagon releasing peptide, vasoactive intestinal
peptide, pituitary adenylated cyclase activating peptide, motilin,
substance P, neuropeptide Y and TSH or an analogue or a fragment
thereof.
63. Microparticles according to claim 62 wherein said drug is
somatostatin or LHRH or an analogue or a fragment thereof.
64. Microparticles according to claim 63 wherein said somatostatin
analogue is D-.beta.-Nal-c[Cys-Tyr-D-Trp-Val-Cys]-Thr-NH.sub.2.
65. A method according to claim 21 wherein said polyester comprises
lactic acid, glycolic acid and tartaric acid.
66. A method according to claim 28 wherein said polyester comprises
lactic acid, glycolic acid and tartaric acid.
67. A method according to claim 37 wherein said polyester comprises
lactic acid, glycolic acid and tartaric acid.
Description
DESCRIPTION
Sustained release ionic conjugate
[0001] 1. Technical Field
[0002] This invention relates to sustained release drug delivery
systems and, in particular, to a method of making microparticles of
a sustained release ionic conjugate.
[0003] 2. Background Art
[0004] Biodegradable polymeric drug delivery formulations have been
developed and utilized for the controlled in vivo release of drugs.
See e.g., U.S. Pat. Nos. 3,773,919 and 4,767,628. Such
biodegradable polymeric formulations are designed to allow an
entrapped drug to slowly diffuse through a polymer matrix or
coating when the biodegradable polymer is depolymerized.
[0005] International Publication No. WO 94/15587 describes
sustained release ionic molecular conjugates of polyesters and
drugs. Since polyester degradation is a key step in the release
process, the surface area of the conjugate particles can control
the release profile of the drug from the conjugate. Thus, the
conjugate particles should be of similar size and shape to insure
both the minimum and reproducible surface area, e.g.,
microspheres.
DISCLOSURE OF INVENTION
[0006] In one aspect, this invention features a method of making
microparticles of a sustained release ionic conjugate containing a
free carboxyl group-containing biodegradable polymer (a polyester
made of monomers such as lactic acid, e-caprolic acid, glycolic
acid, trimethylene carbonate, or p-dioxanone; or a copolymer
thereof; the monomers can be optically isomers or racemates) and a
free amino group-containing drug (e.g., a peptide drug such as
somatostatin or LHRH) which are ionically bonded to each other. The
method includes the steps of (1) obtaining a first solution in
which the conjugate is dissolved; (2) mixing the first solution
(added as small droplets, e.g., through an atomizing nozzle such as
sonic nozzle, pneumatic nozzle, rotary atomizer, or pressure
nozzle) with a first liquid to form a first dispersion, wherein the
first liquid is miscible with the first solution, and the conjugate
is not soluble in the first liquid and precipitates out of the
first dispersion; and (3) isolating the conjugate from the first
dispersion.
[0007] In one embodiment, the drug is soluble in the first liquid,
which can be an alcohol (e.g., ethanol or isopropyl alcohol),
hexane, or water; or a mixture thereof. When ethanol is used as the
first liquid, it can be maintained between about 0.degree. C. and
-30.degree. C. when it is being used. When isopropyl alcohol is
used, it can be maintained between about 0.degree. C. and
-70.degree. C., e.g., cooled by the addition of dry ice.
[0008] The first solution, which may contain acetone,
dichloromethane, acetonitrile, ethyl acetate, tetrahydrofuran, or
glyme, or mixtures thereof can be obtained by (1) dissolving the
biodegradable polymer in a second liquid (e.g., acetone,
tetrahydrofuran, glycone, ethyl acetate, methyl acetate,
acetonitrile, ethyl formate, or glyme; or a mixture thereof) to
form a second solution; (2) dissolving the drug in a third liquid
(e.g., water or acetone; or a mixture thereof) to form a third
solution, wherein the third liquid is miscible with the first
liquid and the second liquid; and (3) mixing the second solution
and the third solution to form the first solution, wherein the
mixing causes the drug to ionically bond to the biodegradable
polymer and form the conjugate in the first solution. The first
solution may comprise up to 40% by weight of the conjugate (e.g.,
between 25 and 35 percent by weight of the conjugate). In one
example, a base, e.g., NaOH or KOH, can be added to the second
solution prior to mixing the second solution and the third
solution. Neutralization of the carboxyl groups of the
biodegradable polymer with the base facilitates the formation of
the ionic conjugate.
[0009] Alternatively, the first solution is obtained by dissolving
the biodegradable polymer and the drug in a second liquid (e.g.,
acetone or a mixture of acetone and water) to form the first
solution, thereby forming the conjugate in the first solution.
According to this method, the biodegradable polymer can be first
dissolved in the second liquid, a base is then added to the second
solution, and the drug is subsequently dissolved in the second
liquid. Also, if desired, the first solution can be partially or
completely evaporated from the first dispersion prior to isolation
of the conjugate. The processed conjugate can be conveniently
isolated by centrifuging or filtering the first dispersion, and the
isolated conjugate can be mixed with an aqueous mannitol solution
prior to vacuum drying (e.g., lyophilization). The isolated
conjugate can be further shaped into a film or a rod. The isolated
conjugate can also be spherified into microspheres of average
diameter of 5 to 200 .mu.m, e.g., as described herein. By
"spherification" or "spherifying" is meant the processing of a
microparticle into a shape close to a sphere.
[0010] In another aspect, this invention features a method of
spherifying a sustained release ionic conjugate as described above.
The method includes the steps of (1) mixing the conjugate with a
first liquid (e.g., an oil such as silicon oil, mineral oil, sesame
oil, or a vegetable oil) to form a first dispersion, wherein the
conjugate has the shape of a microparticle and is not soluble in
the first liquid; (2) heating the first dispersion to a temperature
greater than the Tg or Tm of the conjugate; (3) cooling the first
dispersion below the Tg or Tm of the conjugate; (4) mixing the
first dispersion with a second liquid (e.g., hexane, heptane,
isopropyl myristate, or an alcohol such as ethanol or isopropyl
alcohol) to form a second dispersion, wherein the second liquid is
miscible with the first liquid and the conjugate is not soluble in
the second liquid; and (5) isolating the conjugate from the second
dispersion. The conjugate may have the shape of a microcapsule with
an average diameter of between 5 .mu.m to 200 .mu.m prior to mixing
with the first liquid, and the first dispersion thus formed is
vigorously stirred while being heated to aid in the separation of
the particles. Once the conjugate has been isolated, it can be
rinsed with the second liquid and then vacuum dried. Optionally, it
can also be mixed with an aqueous mannitol solution prior to vacuum
drying.
[0011] A third aspect of this invention features a method of
spherifying the above-described sustained release ionic conjugate
(e.g., a microcapsule having an average diameter of between 5 .mu.m
to 200 .mu.m). The method includes the steps of (1) mixing the
conjugate in a first liquid (e.g., water) to form a first
dispersion, wherein the conjugate is in the shape of a
microparticle and the conjugate is not soluble in the first liquid;
(2) stirring the first dispersion; (3) mixing the stirred
dispersion with a second liquid (e.g., dichloromethane or
chloroform) in such an amount so that it is absorbed by the
conjugate but does not solubilize the conjugate, wherein the second
liquid is miscible with the first liquid; (4) evaporating the
second liquid from the first dispersion; and (5) isolating the
precipitated conjugate from the first dispersion. If necessary, the
method may further include the step of adding a surfactant (e.g.,
lecithin, Tween 20, polysorbate, or lauryl sulfate) to the first
dispersion to aid in the stabilization of the first dispersion, and
the isolated conjugate can be rinsed with the first liquid and
vacuum dried. Again, the isolated conjugate can be mixed with an
aqueous mannitol solution prior to vacuum drying.
[0012] In a further aspect of this invention, this invention
features a method of spherifying the above-described sustained
release ionic conjugate. The method includes the steps of (1)
dissolving the conjugate in a first liquid (e.g., acetonitrile) to
form a first solution; (2) stirring the first solution with a
second liquid (e.g., an oil) to form a first dispersion, wherein
the second liquid is immiscible with the first solution; (3)
evaporating the first liquid from the first dispersion to
precipitate the conjugate from the first dispersion; and (4)
isolating the precipitate conjugate from the first dispersion. In
the stirring step, the first solution can be added to the second
liquid as small droplets.
[0013] The above method can further include the step of rinsing the
isolated conjugate with a third liquid (e.g., hexane, heptane, or
octane) which is miscible with the second liquid and not a solvent
for the isolated conjugate. If desired, the isolated conjugate can
be mixed with an aqueous mannitol solution prior to vacuum
drying.
[0014] The biodegradable polymer in the above-described conjugate
may contain at least one free carboxyl group (e.g., two to ten free
carboxyl groups per polymer chain). Examples of carboxylic acid
containing biodegradable polymers include polyesters containing
units of lactic acid, e-caprolic acid, p-dioxanone, e-caprionic
acid, substituted and unsubstituted trimethylene carbonate,
1,5-dioxepan-2-one, 1,4-dioxepan-2-one, glycolic acid, alkylene
oxylate, cycloalkylene, cycloalkylene oxylate, alkylene succinate,
or 3-hydroxy butyrate in optically active forms or as racemates; or
copolymers of any of the above. Additional free carboxylic acid
groups can be incorporated into the biodegradable polyester by
reaction, e.g., ring opening polymerization or polycondensation,
with polycarboxylic acids such as malic acid, tartaric acid, pamoic
acid, citric acid, succinic anhydride, and glutaric anhydride.
Thus, the biodegradable polymer can be a water insoluble polyester
including lactic acid units with or without glycolic acid units.
Other biodegradable polymers such as polyorthoesters,
polyorthocarbonates, and polyantals may also be used. The
biodegradable polymer may have an average degree of polymerization,
e.g., average number of monomers per polymer chain, between 10 and
300.
[0015] The drug has one or more (e.g., one to ten) free amine
groups. In one embodiment, the drug is an acid-stable peptide.
Examples of suitable acid-stable peptides include growth hormone
releasing peptide (GHRP), luteinizing hormone-releasing hormone
(LHRH), adrenomedullin, growth hormone, somatostatin, bombesin,
gastrin releasing peptide (GRP), calcitonin, bradykinin, galanin,
melanocyte stimulating hormone (MSH), growth hormone releasing
factor (GRF), amylin, adrenomedullin, tachykinins, secretin,
parathyroid hormone (PTH), enkephalin, endothelin, calcitonin gene
releasing peptide (CGRP), neuromedins, parathyroid hormone related
protein (PTHrP), glucagon, neurotensin, adrenocorticotrophic
hormone (ACTH), peptide YY (PYY), glucagon releasing peptide (GLP),
vasoactive intestinal peptide (VIP), pituitary adenylated cyclase
activating peptide (PACAP), motilin, substance P, neuropeptide Y
(NPY), TSH, and analogs and fragments thereof. The drug may be
soluble (e.g., greater than 0.1 mg/ml; preferably, greater than 1.0
mg/ml) in the first liquid.
[0016] Other features and advantages of the present invention will
be apparent from the detailed description and from the claims.
[0017] It is believed that one skilled in the art can, based on the
description herein, utilize the present invention to its fullest
extent. The following specific embodiments are, therefore, to be
construed as merely illustrative, and not limitative of the
remainder of the disclosure in any way whatsoever.
[0018] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as conmnonly understood by one of
ordinary skill in the art to which this invention belongs. Also,
all publications, patent applications, patents, and other
references mentioned herein are incorporated by reference.
[0019] Best Modes for Carrying Out the Invention
EXAMPLE 1
[0020] 18.0 g of a 6,000 g/mol 66/32/2
poly-L-lactic-co-glycolic-co-D,L-ma- lic acid copolymer (66 percent
L-lactic acid, 32 percent glycolic, and 2 percent malic acid; acid
number of 0.373 milliequivalents/g) was dissolved in 180 g of
acetone (solution of 10% copolymer by weight). 14.4 ml of 0.5 N
aqueous NaOH was added to form the sodium carboxylate of the
polymer. 4.28 g of the acetate salt of the peptide Lanreotide.TM.
(Kinerton, Dublin, Ireland;
D-Nal-c[Cys-Tyr-D-Trp-Val-Cys]-Thr-NH.sub.2; acetate content=9.60
percent by weight) was separately dissolved in a mixture of 10 g of
acetone and 10 g of deionized water. The amount of peptide
dissolved corresponded to the stoichiometric ratio of acid groups
from the copolymer (e.g., one) and the free amino groups for the
peptide (e.g., two). The peptide solution was then added dropwise
to the copolymer solution, and the resulting solution was stirred
for two hours to allow for salt exchange and the resulting
formation of the polymer/peptide ionic conjugate (PPIC).
EXAMPLE 2
[0021] In a temperature controlled jacketed reactor (Schott Glass
AGB, Dublin, Ireland), a two liter bath of deionized water was
precooled to 0.degree. C. and was vigorously stirred. The above
PPIC solution of Example 1 was then slowly added to the reactor
using a Masterflex pump (Bioblock Scientific, Illkvch, France)
which produced a flow rate of 10-15 ml/min through a silicone
tubing fitted with a 19 gauge needle at its tip. The PPIC solution
was fed through the needle that was positioned above a water bath
at 0.degree. C. The PPIC precipitated in the bath as small, solid
particles. The solid particles were then separated from the
supernatant by centrifugation (30 minutes at 5000 rpm and
0-5.degree. C.), rinsed with fresh deionized water, resuspended
water, recentrifuged, and then lyophilized. The isolated conjugate
was filtered through a 100 .mu.m sieve to remove any large
particles which would not be capable of being injected through a 21
gauge needle. An analysis of the resulting particle sizes is
described in Table I.
EXAMPLE 3
[0022] The PPIC solution of Example 1 was also precipitated as
described above in Example 2, except that a bath of ethanol at a
temperature of -20.degree. C. was used instead of a bath of water
at 0.degree. C. An analysis of the resulting particle sizes is
described in Table I.
EXAMPLE 4
[0023] The PPIC solution of Example 1 was also dispersed at a
controlled flow rate of 4 mI/min through an atomizing nozzle
containing a hollow tip (Bioblock; 50 Watts, 20 kHz) over a bath of
ethanol at -10.degree. C. in a temperature controlled jacket
reactor. In this nebulization process, the copolymer solution was
released from the probe as a fine mist of small droplets. The small
droplets fell into the ethanol bath, causing the deionized water
and acetone to be extracted from the droplets. As a result, the
copolymer droplets hardened as small, solid particles. The
particles were then recovered by centrifugation and lyophilized. An
analysis of the resulting particle sizes is listed in Table I. What
is meant by Diameter -10 (i.e., D0.1), Diameter -50 (i.e., D0.5),
or Diameter -90 (i.e., D0.9) is the smallest diameter which is
greater than 10%, 50%, and 90% of the total particles,
respectively. What is meant by specific area is the average
specific area of the resulting particles.
1TABLE I Specific Diameter-10 Diameter-50 Diameter-90 Area Example
(.mu.m) (.mu.m) (.mu.m) (m.sup.2/g) 2 10 30 62 18.64 3 9 37 89 6.42
4 13 46 95 22.61
EXAMPLE 5
[0024] 5.0 g of the PPIC described above in Example 4 was dissolved
in 20 g of acetone (concentration of 20% PPIC by weight). This
solution was then nebulized at a flow rate of 4.0 ml/min over 500
ml of bath of ethanol at -10.degree. C. as described in Example 4.
After preparation of the PPIC particles in the bath, 500 ml of
deionized water was added to the bath, and the bath was then
brought up to 0.degree. C. The bath was then stirred for 30 min,
brought up to 20.degree. C., and stirred for an additional 30 min.
The PPIC particles were then recovered by filtration and dried
under vacuum at room temperature. An analysis of the resulting
particles is listed in Table II.
2TABLE II Specific D 0.1 D 0.5 D 0.9 Area Example (.mu.m) (.mu.m)
(.mu.m) (m.sup.2/g) #4 13 46 95 22.61 #5 50 99 180 0.11
[0025] As shown in Table II, different morphologies of particles
were obtained. The particles of Example 4 were larger and had a
lower specific area. As indicated by electron scanning microscope,
the particles obtained in Example 4 were also more porous, likely
because of the frozen water which remained in the particles upon
precipitation. When the bath dispersion was returned to room
temperature, the water thawed and flowed into the ethanol bath,
leaving open channels in the microparticles. Consequently, these
particles were more brittle and generated fragments of small
size.
EXAMPLE 6
[0026] A solution of PPIC described above in Example 5 was
nebulized at 2.5 mmin over 1.5 liters of deionized water at
0.degree. C. An analysis of the resulting particle sizes is listed
in Table III.
EXAMPLE 7
[0027] A solution of PPIC described above in Example 5 was
nebulized at 2.5 m/min over 1.5 liters of ethanol at -10.degree. C.
An analysis of the resulting particle sizes is listed in Table
III.
3TABLE III Specific D 0.1 D 0.5 D 0.9 Area Example (.mu.m) (.mu.m)
(.mu.m) (m.sup.2/g) 6 53.4 154.3 329.1 n/a 7 42.4 87.2 170.1
0.20
EXAMPLE 8
[0028] Two solutions of PPIC are prepared in acetone as described
above in Example 5. The first solution has a PPIC concentration of
15% while the second solution has a PPIC concentration of 20%. The
solutions are nebulized over a bath of ethanol at -10.degree. C. at
flow rates of 2.5, 3.5, and 5.0 mmin as described in Example 5. An
analysis of the resulting particle size is listed in Table IV.
4TABLE IV Specific Feeding Rate D 0.1 D 0.5 D 0.9 Area
Concentration (ml/min) (.mu.m) (.mu.m) (.mu.m) (m.sup.2/g) 15% 2.5
35.9 81.6 191.1 4.455 15% 3.5 34.4 80.2 188.3 8.336 15% 5.0 49.4
163.6 397.8 n/a 20% 2.5 33.3 73.8 145.6 0.199 20% 3.5 50.8 112.7
241.9 0.579 20% 5.0 108.3 219.1 395.9 n/a
[0029] Analysis of the particles using scanning electron microscope
revealed that the particle size and the specific area increased
with the increase in feeding rate.
EXAMPLE 9
[0030] 5.0 g of PPIC microparticles of Example 4 was dissolved in
45 g of acetone (concentration 10% by weight). The solution was
then added dropwise over a vigorously stirred 500 ml n-hexane at
room temperature. The n-hexane solution turned cloudy as particles
of PPIC precipitated. The PPIC was removed by filtration and dried
under vacuum at room temperature.
EXAMPLE 10
[0031] In a jacketed reactor, 3.0 g of the PPIC microparticles
described in Example 2 was dispersed in a vigorously stirred 250 ml
of 12,500 cs medical grade silicon oil (Dow Coming, Midland, Mich.)
(of 1% PPIC by weight). After the stirring, the mixture was then
heated to 120.degree. C., which is above the Tg of 55.degree. C.
for the PPIC, and kept at this temperature for 30 minutes. During
this heating, the isolated individual particles melted to form
spherical droplets. The dispersion was then cooled to 20.degree. C.
and then diluted with 1250 ml of hexane. The microspheres
subsequently hardened, were recovered by filtration, were rinsed
with fresh hexane, and were finally dried under vacuum. The
characteristics of the obtained microspheres are reported in Table
V. The final microspheres had a small diameter as compared to those
of Example 2 as a result of the compaction of the particles during
melting.
5TABLE V Specific D 0.1 D 0.5 D 0.9 Area Example (.mu.m) (.mu.m)
(.mu.m) (m.sup.2/g) #2 10 30 62 18.64 #7 2 10 47 <0.33
Example 11
[0032] 0.2 g of the PPIC microparticles described in Example 2 were
dispersed in 5 ml of deionized water and vigorously stirred with a
vortex shaker. 100 ml microliters of dichloromethane (DCM) was then
added over the stirred dispersion. The addition of a small amount
of DCM caused a swelling of the surface of the PPIC particles.
Stirring was kept at room temperature for 4 hours, allowing for
evaporation of DCM and consequential hardening of the swollen
surface of the particles. A scanning electron microscope showed
that the resulting particles were of spherical shape with a
smoother surface as compared to the starting material. Both the
particle size distribution was narrowed, and the maximum particle
size was reduced as a result of the increase in the density of the
particles.
EXAMPLE 12
[0033] One liter of sesame seed oil (Vitamins, Inc., Chicago, Ill.)
was placed in a 2 liter, three necked flask immersed in a water
bath. The oil was stirred at 600 rpm using a teflon stirring paddle
connected to an overhead stirring motor. 500 mg of the surfactant,
soybean lecithin, (Sigma Chemicals, St. Louis, Mo.) was added to
the sesame seed oil, and the mixture was stirred for 10 min. 10 g
of a PPIC formulation was then dissolved in 100 ml acetonitrile to
give a clear solution. The PPIC compositions were made using
Lanreotide.TM. conjugated with one of the following three polymers:
64/34/2 poly-DL-lactic-co-glycolic-D,L-malic acid copolymer (M. W.
avg 6,000) (Composition 1); 74/24/2
poly-DL-lactic-co-glycolic-D,L-malic acid copolymer (M.W. avg
6,000) (Composition 2); and 98/2 poly-DL-lactic-co-D,L-malic acid
copolymer (Composition 3).
[0034] This clear PPIC solution was added dropwise through a
dropping funnel. When the addition was completed, the temperature
of the external water bath was raised to 40.degree. C., and the oil
was left stirring for 20 h. One liter of hexane was then added to
dilute the sesame seed oil, and the oil was filtered through a
medium fritted funnel. The microspheres collected in the filter
funnel were further washed several times with 500 ml in total
volume of hexane. The particles are dried at 36.degree. C. for two
days under vacuum. Characteristics of the resulting microspheres
are presented in Table VI.
6TABLE VI Specific D 0.1 D 0.5 D 0.9 Area Composition (.mu.m)
(.mu.m) (.mu.m) (m.sup.2/g) 1 13 28 57 0.1426 2 13 25 59 0.1395 3
14 25 51 0.1480
EXAMPLE 13
[0035] Reactor was loaded with monomers glycolide (Purac Biochem,
Netherlands, 84.83 g), lactide (Purac Biochem, Netherlands, 210.67
g) and L(+)-tartaric acid (Riedel-de Haen, Seelze, Germany, article
number 33,801, 4.50 g) and stannous 2-ethyl hexanoate (Sigma, St.
Louis, Mo., USA, article number S-3252) in toluene (Riedel-de Haen,
Seelze, Germany) solution (0.1025 M, 4.34 ml). The L(+)-tartaric
acid was previously dried over phosphorous pentoxide (Riedel-de
Haen, Seelze, Germany) in an Abderhalden drying apparatus for 10
hours. The reactor (connected to pump via a liquid nitrogen trap)
was then put under vacuum (0.04 mbar) with stirring for 50 minutes
to remove toluene. The reactor, under an atmosphere of oxygen-free
nitrogen (BOC gases, Dublin, Ireland, moisture content of 8 VPM),
was then immersed in an oil bath (Temperature=200.degree. C.) and
stirring was increased to 125 rpm. Prior to immersion, a heating
tape (Thermolyne type 45500, input control setting=4) was placed on
the reactor lid. The time taken to completely melt the reactor
contents was noted, typically 10 minutes for a load of 300 g at
200.degree. C. Samples were taken every hour during synthesis and
analyzed by GPC to determine the percentage residual monomer and to
obtain values for average molecular weight by number (Mn) and by
weight (Mw). Typical reaction times are of the order of 6
hours.
[0036] An amorphous copolymer was obtained comprising 66.21%
lactide units, 33.11% glycolide units, and 0.68% tartaric acid
units (66/33/1 PLGTA). The acid number of the titration was
determined to 0.303 milliequivalents/g (meq/g; the normality of
NaOH multiplied by the volume of NaOH solution required to
neutralize one gram of polyester). The average number average
molecular weight of the copolymer had a value of 10,250, the
average weight molecular weight of the copolymer was 11,910 giving
a Mw/Mn value of 1.16.
[0037] 41.32 g of the above 10,000 g/mol 66/32/2
poly-L-lactic-co-glycolic- -co-L(+)-tartaric acid copolymer (acid
number=0.303 meq/g) was dissolved in 165.52 g of acetone (Riedel-de
Haen, Seelze, Germany) by sonication in a Branson sonication bath
(Branson, Danbury, Conn., USA) to give a solution with a
concentration of 19.98% PLGTA by weight.
[0038] To this solution was added 37.6 ml 0.2 N sodium carbonate
(Aldrich, Gillingham, Dorset, UK) thus providing a 1.2 times excess
of sodium over copolymer carboxyl groups. The solution was left to
stir for 30 minutes to aid sodium salt formation. It was then fed
to an atomizer nozzle at 8.0 ml/min using a Masterflex pump (Cole
Parmer, Barrington, Ill., USA). The solution was nebulized into a 6
L jacketed reactor containing 2 L of de-ionized water cooled to
2.5.degree. C. using a circulation bath (Huber, Offenburg,
Germany). This water was stirred to 350 rpm using a 4-blade paddle
linked to a stirrer motor.
[0039] Once nebulization was complete, the dispersion was placed in
6 centrifuge bottles and spun at 5000 rpm for 30 minutes in a
Sorvall centrifuge (DuPont Sorvall Products, Wilmington, Del.,
USA). The resultant centrifuge cakes were resuspended in de-ionized
water and re-spun. The supematant was discarded and the cakes were
frozen in a freezer overnight before being dried in a small-scale
lyophilizer (Edwards, Crawley, West Sussex, UK) the next day. 33.16
g of washed copolymer were recovered representing a yield of
80.24%.
[0040] 4.92 g of the above 10,000 g/mol 66/33/1
poly-L-lactic-co-glycolic-- co-D,L-tartaric acid copolymer (66
percent L-lactic acid, 33 percent glycolic acid, and 1 percent
tartaric acid) was dissolved in 11.58 g of acetonitrile (Ridel-de
Haen, Seelze, Germany; HPLC grade) by sonication in a Branson
sonication bath (Branson, Danbury, CT, USA) and stirred on a
stir-plate producing a solution with a concentration of 29.82% of
PLGTA by weight.
[0041] This copolymer/acetonitrile solution was fed from a glass
reservoir through an atomizer nozzle using an FMI revolving piston
pump (FMI, Oyster Bay, N.Y., USA) set at 2.0 ml/min. The atomizer
output power was set at 50 W with an amplitude of 80%. The solution
was nebulized into a 6L jacketed reactor containing 1.5 L of
general purpose reagent isopropyl alcohol (Labscan, Dublin,
Ireland), cooled to -70.degree. C. by solid CO.sub.2 pellets (AIG,
Dublin, Ireland), and stirred at 300 rpm with a 4-blade paddle
linked to a stirrer motor. The temperature of the isopropyl alcohol
remained at or close to -70.degree. C. throughout nebulization
which lasted for approximately 8 minutes.
[0042] Once nebulization was complete, the dispersion was allowed
to warm to 10.degree. C. of its own accord over a period of 5.5
hours. It was then filtered over a Whatman No. 1 filter paper (9 cm
diameter) with the aid of a vacuum. The filter paper and cake were
placed in a desiccator along with silica gel drying chips and a
vacuum was pulled through an automatic refrigeration trap at
-110.degree. C. After 24 hours 4.24 g of material were recovered.
An analysis of the resulting particles is listed in Table VII.
7TABLE VII Example D 0.1 D 0.5 D 0.9 Specific Area # (pm) (pm) (pm)
(.mu..sup.2/g) 13 31 68 139 0.16
[0043] It is to be understood that while the invention has been
described in conjunction with the detailed description thereof,
that the foregoing description is intended to illustrate and not
limit the scope of the invention, which is defined by the scope of
the appended claims. Other aspects, advantages, and modifications
are within the claims.
* * * * *